One question regarding PDEs relates to cell-specific expression, regulation and function of specific PDE isoforms. Distinct distributions of rat PD3A and B mRNAs were observed in Northern Blots and in situ hybridization. PDE3 isoforms have a limited distribution in human immune/inflammatory cells. cDNA cloning (human lymphocyte cDNA library) and RT-PCR amplification identified PDE3B, not PDE3A, mRNA in T- lymphocytes and peripheral macrophages. T-lymphocytes and macrophages also contain PDE4. Individual T-lymphocyte clones sensitized to basic myelin protein contain different proportions of PDE3 and PDE4 activities (measured with 0.1 uM cAMP) which increase during antigen- stimulated proliferation. The proliferative response is inhibited by specific inhibitors of PDE3 (cilostamide) and PDE4 (rolipram), consistent with the known effects of cAMP on immune/inflammatory cell functions. PDE3 activity was much lower relative to that of PDE4 in EB virus transformed beta-lymphocytes than in T-lymphocytes, NK cells and macrophages. Other workers have reported little or no PDE3 activity in polymorphonuclear leukocytes, eosinophils, basophils, monocytes. In peripheral macrophages PDE3 activity > PDE4; in two mononuclear cell lines, PDE3 > 4 in THP-1 cells and PDE4 >> PDE3 in U937 cells. It is possible that PDE3 activity increases during monocyte/macrophage differentiation. PDE3A isoforms are present in platelets and HEL erythroleukemia cells; by in situ hybridization, rat PDE3A, not B, mRNA was detected in megakaryocytes present in developing liver. The mechanisms for and functional consequences of this differential distribution/expression of PDE3A and B isoforms in immune/inflammatory cells and megakaryocytes/platelets is not known. Understanding of cellular regulation of specific PDE isoforms will be of increasing importance for understanding pathophysiology and for targeting specific PDEs in treatment of pulmonary disorders, especially those relating to inflammation, allergy, and airway and vascular reactivity. To learn more about PDE3B function both during development and in differentiated cells, we are attempting to "knockout" the mouse PDE3B by homologous recombination.